Transfection during cell synchronisation (to G2/M)
I'd like to transiently overexpress a plasmid encoded gene in HeLa and be able to detect the protein after synchronisation to G2/M. I've tried to transfect with Lipofectamine in the approx 3h window between the Thymidine (S phase) and Nocodazole (G2/M) blocks, but had no overexpression (unsync cells were transfected well) and suspect the cells may be refractory until they can enter mitosis (Mortimer, I. Gene Therapy (1999)) I suspect I will run into problems with cell over confluency if I seed to 50-70% for transfection before cell sync. Please can you offer some advice. Thanks.